The long-term objective of proposed research is to identify and characterize those steps in viral RNA biosynthesis at which virus expression in cytomegalovirus (CMV) infected cells is regulated. One goal of the studies proposed is to characterize the biosynthesis of m-RNAs encoded in specific regions of the CMV genome. The specific aims are to determine the size classes, map positions, direction of transcription, translation products, and, if present, the map positions of splice points of m-RNAs encoded in the repeat and repeat-unique-joint regions of the CMV Towne genome. A second goal is to characterize nucleotide sequence heterogeneity observed among strains of CMV. The specific aims are to map more precisely the positions of sequence heterogeneity between the DNAs of the Towne and AD169 strains of CMV, and to determine whether the sequences unique to each strain can be used as markers to provide a basis for placing CMV isolates into groups. Cloned restriction fragments representing the repeat-unique-joint regions will be bound to cellulose and used to select cytoplasmic polyadenylated RNA. To determine sizes and map positions, the selected RNA will be size separated in methylmercury agarose gels, transferred to nitrocellulose, and hybridized with the appropriate in vitro labeled restriction fragments. To detect splice points, the selected RNAs will be hybridized with the appropriate restriction fragments and the hybrids then analyzed by the nuclease S1 mapping technique. The direction of transcription will be determined by labeling the 5 foot ends of DNA fragments prior to nuclease S1 mapping. The repeat and repeat-unique-joint regions are those regions where nucleotide sequence deletions and sequence heterogeneity between CMV strains have been mapped. The functional significance of these regions of the genome would be established as a result of these studies. Also, the possible etiological and epidemiological implications of strain unique sequences in these regions of the genome are clinically relevant.